Ligia Maria Oliveira Chueire

The complete genome sequence of Chromobacterium violaceum reveals remarkable and exploitable bacterial adaptability

Chromobacterium violaceum is one of millions of species of free-living microorganisms that populate the soil and water in the extant areas of tropical biodiversity around the world. Its complete genome sequence reveals (i) extensive alternative pathways for energy generation, (ii) ≈500 ORFs for transport-related proteins, (iii) complex and extensive systems for stress adaptation and motility, and (iv) widespread utilization of quorum sensing for control of inducible systems, all of which underpin the versatility and adaptability of the organism. The genome also contains extensive but incomplete arrays of ORFs coding for proteins associated with mammalian pathogenicity, possibly involved in the occasional but often fatal cases of human C. violaceum infection. There is, in addition, a series of previously unknown but important enzymes and secondary metabolites including paraquat-inducible proteins, drug and heavy-metal-resistance proteins, multiple chitinases, and proteins for the detoxification of xenobiotics that may have biotechnological applications.

Isolation and characterization of new efficient and competitive bean (Phaseolus vulgaris L.) rhizobia from Brazil

The common bean (Phaseolus vulgaris L.) is widely cultivated in South and Central America and Africa, but inoculation with rhizobia often does not lead to a response in field experiments. A selection program was started in the State of Parana, Brazil, in which three promising strains, PRF 35, PRF 54 and PRF 81, showing high rates of N2 fixation, were competitive and tolerated high temperatures. The performance of the strains was also verified in four field experiments, where inoculation with PRF 81 allowed yield increases of up to 906 kg ha−1, compared with the non-inoculated (control) with a high population of native bean rhizobia. The high performance of PRF 81 was confirmed in several other field trials carried out in Brazil, leading to its recommendation for use in commercial Brazilian inoculants. PRF 34, PRF 54 and PRF 81 were further characterized and compared with four strains, representative of bean rhizobia species in an effort to define variables which could aid future selection programs. The Brazilian strains showed unique profiles of protein, lipopolysaccharide and PCR using specific (ERIC and REP) or arbitrary short primers. The DNA fingerprints obtained with specific or arbitrary primers showed that strains PRF 35 and PRF 54 were genetically very close, nevertheless, there were substantial differences between the strains in nodulation and N2 fixation rates, as well as in the synthesis of Nod factors after induction with naringenin. The Brazilian strains showed Nod factor profiles similar to those of R. tropici type IIA CFN 299 and IIB CIAT 899 strains, and mixed characteristics of both types. That is, they were unable to grow in LB and PY minus Ca, as with type IIA, but were tolerant to high temperature, acidity, and had the same PCR product with Y1 and Y2 primers, as type IIB strain. The Brazilian strains showed mixed host range spectra between strain types IIA and IIB and, by the analysis of 17 fatty acids, strains PRF 35 and PRF 54 were grouped with CFN 299 and PRF 81 with CIAT 899. The performance of strain PRF 81 in field experiments indicates future potential for identification of new competitive and efficient R. tropici strains for tropical and subtropical areas.

Genome of Herbaspirillum seropedicae Strain SmR1, a Specialized Diazotrophic Endophyte of Tropical Grasses

The molecular mechanisms of plant recognition, colonization, and nutrient exchange between diazotrophic endophytes and plants are scarcely known. Herbaspirillum seropedicae is an endophytic bacterium capable of colonizing intercellular spaces of grasses such as rice and sugar cane. The genome of H. seropedicae strain SmR1 was sequenced and annotated by The Paraná State Genome Programme—GENOPAR. The genome is composed of a circular chromosome of 5,513,887 bp and contains a total of 4,804 genes. The genome sequence revealed that H. seropedicae is a highly versatile microorganism with capacity to metabolize a wide range of carbon and nitrogen sources and with possession of four distinct terminal oxidases. The genome contains a multitude of protein secretion systems, including type I, type II, type III, type V, and type VI secretion systems, and type IV pili, suggesting a high potential to interact with host plants. H. seropedicae is able to synthesize indole acetic acid as reflected by the four IAA biosynthetic pathways present. A gene coding for ACC deaminase, which may be involved in modulating the associated plant ethylene-signaling pathway, is also present. Genes for hemagglutinins/hemolysins/adhesins were found and may play a role in plant cell surface adhesion. These features may endow H. seropedicae with the ability to establish an endophytic life-style in a large number of plant species.

Tillage method and crop rotation effects on the population sizes and diversity of bradyrhizobia nodulating soybean.

Abstract This study was conducted in an area of Brazil cultivated with soybean since the early 1960s but which for the last 17 yr was under different tillage (no-tillage, NT; conventional tillage, CT) and crop rotation (soybean, S/wheat, W/maize, M; S/W; M/W) systems. The area had not received any inoculant for the last 15 yr and our objective was to investigate the effects of tillage and cropping systems on the bradyrhizobia population. The NT system and crop rotations with soybean resulted in high populations of bradyrhizobia, but even in the treatment where soybean had not been cultivated for 17 yr (M/W) the number of viable cells in the soil was high. A total of 142 bradyrhizobia isolated from the different treatments were characterized based on colony morphology, serological reaction, DNA analysis by RAPD, protein and Nod factors profiles. The analyses resulted in grouping of the isolates into 16 DNA, five protein and three Nod factors profiles. A high proportion (37.5%) of the isolates did not react with any known serogroup. Both NT and crop rotations with soybean resulted in a higher bradyrhizobia diversity, with the lowest number of genomic patterns occurring in the CT with M/W rotation. However, there was no relationship between the treatment combinations and genetic relatedness. The evaluation of symbiotic performance under greenhouse conditions showed that the isolates with higher rates of N2 fixation were also isolated from NT with S/W or S/W/M crop rotations. Consequently, the use of agronomic practices such as NT and crop rotation with legumes will not only contribute to agricultural sustainability, but also help to maintain bradyrhizobia population and diversity.

Genomic basis of broad host range and environmental adaptability of Rhizobium tropici CIAT 899 and Rhizobium sp. PRF 81 which are used in inoculants for common bean (Phaseolus vulgaris L.).

BackgroundRhizobium tropici CIAT 899 and Rhizobium sp. PRF 81 are α-Proteobacteria that establish nitrogen-fixing symbioses with a range of legume hosts. These strains are broadly used in commercial inoculants for application to common bean (Phaseolus vulgaris) in South America and Africa. Both strains display intrinsic resistance to several abiotic stressful conditions such as low soil pH and high temperatures, which are common in tropical environments, and to several antimicrobials, including pesticides. The genetic determinants of these interesting characteristics remain largely unknown.ResultsGenome sequencing revealed that CIAT 899 and PRF 81 share a highly-conserved symbiotic plasmid (pSym) that is present also in Rhizobium leucaenae CFN 299, a rhizobium displaying a similar host range. This pSym seems to have arisen by a co-integration event between two replicons. Remarkably, three distinct nodA genes were found in the pSym, a characteristic that may contribute to the broad host range of these rhizobia. Genes for biosynthesis and modulation of plant-hormone levels were also identified in the pSym. Analysis of genes involved in stress response showed that CIAT 899 and PRF 81 are well equipped to cope with low pH, high temperatures and also with oxidative and osmotic stresses. Interestingly, the genomes of CIAT 899 and PRF 81 had large numbers of genes encoding drug-efflux systems, which may explain their high resistance to antimicrobials. Genome analysis also revealed a wide array of traits that may allow these strains to be successful rhizosphere colonizers, including surface polysaccharides, uptake transporters and catabolic enzymes for nutrients, diverse iron-acquisition systems, cell wall-degrading enzymes, type I and IV pili, and novel T1SS and T5SS secreted adhesins.ConclusionsAvailability of the complete genome sequences of CIAT 899 and PRF 81 may be exploited in further efforts to understand the interaction of tropical rhizobia with common bean and other legume hosts.

Genetic diversity of indigenous tropical fast-growing rhizobia isolated from soybean nodules

This study characterized genetically 30 fast-growing rhizobial strains isolated from nodules of Asian and modern soybean genotypes that had been inoculated with soils from disparate regions of Brazil. Analyses by rep-PCR (ERIC and REP) and RAPD indicated a high level of genetic diversity among the strains. The RFLP-PCR and sequencing analysis of the 16S rRNA genes indicated that none of the strains was related to Sinorhizobium (Ensifer) fredii, whereas most were related to Rhizobium tropici (although they were unable to nodulate Phaseolus vulgaris) and to Rhizobium genomic species Q. One strain was related to Rhizobium sp. OR 191, while two others were closely related to Agrobacterium (Rhizobium) spp.; furthermore, symbiotic effectiveness with soybean was maintained in those strains. Five strains were related to Bradyrhizobium japonicum and B. elkanii, with four of them being similar to strains carried in Brazilian inoculants, therefore modifications in physiological properties, as a shorter doubling time might have resulted from adaptation to local conditions. Phospholipid fatty acid analysis (PFLA) was less precise in delineating taxonomic relationships. The strains fit into eight Nod-factor profiles that were related to rhizobial species, but not to N2-fixation capacity or competitiveness. The data obtained highlight the diversity and promiscuity of rhizobia in the tropics, being capable of nodulating exotic legumes and might reflect ecological strategies to survive in N-poor soils; in addition, the diversity could also represent an important source of efficient and competitive rhizobial strains for the tropics. Putative new rhizobial species were detected only in undisturbed soils. Three species (R. tropici, B. japonicum and B. elkanii) were found under the more sustainable management system known as no-till, while the only species isolated from soils under conventional till was R. tropici. Those results emphasize that from the moment that agriculture was introduced into undisturbed soils rhizobial diversity has changed, being drastically reduced when a less sustainable soil management system was adopted.

Genome Sequences of Burkholderia sp. Strains CCGE1002 and H160, Isolated from Legume Nodules in Mexico and Brazil

The genome sequences of Burkholderia sp. strains CCGE1002 from Mexico and H160 from Brazil, isolated from legume nodules, are reported. Their gene contents in relation to plant-microbe interactions and xenobiotic degradation are discussed.

Avaliação qualitativa e quantitativa da microbiota do solo e da fixação biológica do nitrogênio pela soja

The objective oh this work was to define, for edaphoclimatic conditions of Brazil, acceptable levels of dispersion of some biological parameters, used in studies of environmental impact due to the adoption of new technologies with the soybean crop. Two experiments were performed in eleven counties, in six states and in the Federal District, with conventional and transgenic soybean. Evaluated parameters were: carbon and nitrogen of the microbial biomass, microbial respiration, microbial metabolic quotient, electrophoresis of soil DNA in denaturating gradient gel (DGGE), biological nitrogen fixation, number of rhizobial cells, nodule number and nodule dry weight, nodule occupancy by Bradyrhizobium strains, shoot dry weight, total nitrogen in shoot and nitrogen as ureides in shoot. The temporal variability for carbon and nitrogen of microbial biomass, microbial respiration and microbial metabolic quotient was adequate, and the maximum coefficient of variation was estimated at 35%. The homogeneity between replicates, treatments and harvests was confirmed by the DGGE method. In N-poor soils, the parameters of nodules and shoot dry weight, with maximum coefficient of variation 33 and 18%, respectively, were adequate for evaluation of biological nitrogen fixation, which contributed from 72 to 88% of total nitrogen in shoot.

The Brazilian Experience with the Soybean (Glycine max) and Common Bean (Phaseolus vulgaris) Symbioses

Soybean and common bean are cultivated on more than 17 million hectares in Brazil. Soybean is the main cash crop and relies exclusively on BNF to supply the plant N demands. However, to maintain yields of more productive cultivars, a search for efficient, competitive and stress tolerant strains continues. Common bean is the most important legume used for food and inoculation with an efficient, competitive Rhizobium tropici strain increases in nodulation, nodule occupancy and yield. An adequate soil management is essential to guarantee the benefits of BNF with both crops.

Genome Sequence of Rhizobium ecuadorense Strain CNPSo 671T, an Indigenous N2-Fixing Symbiont of the Ecuadorian Common Bean (Phaseolus vulgaris L.) Genetic Pool

ABSTRACT Rhizobium ecuadorense CNPSo 671T was isolated from a common bean nodule in Ecuador. The draft genome brings novelty about indigenous rhizobial species in centers of genetic diversity of the legume.